Cloning, expression and purification of Mycobacterium tuberculosis furB

    JIANG Hong, XUE Ying, GAO Xue, SHI Changª²Hong, BAI Yinª²Lan, ZHANG Hai, LI Yuan, XU Zhiª²Kai

    Department of Microbiology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To obtain Mycobacterium tuberculosis (MTB) furB from H37Rvª²DNA, express efficiently in E. coli and purify the FurB proteins. METHODS: furB was amplified by PCR from the genome of H37Rv and cloned into pGEMª²Tª²Easy vector. After sequenced, furB was recombinated into the expression vector pQEª²80L by restriction enzyme digestion, which was named pQEª²80Lª²furB. The plasmid pQEª²80Lª²furB was transformed into E.coli DH5¦Á and induced by IPTG. The furB expression was analyzed by SDSª²PAGE and confirmed by Western blot with miceª²specific mAb against (His)6. RESULTS: furB was identical to what reported by Genbank. The pQEª²80Lª²furB vector expressed protein with relative molecule mass 15.0¡Á103, which could be caught by (His)6 mAb. The expressed protein could be purified by Niª²NTA system kit with denatured condition. CONCLUSION: furB has been successfully cloned and efficiently expressed in E.coli, The results lay the basis for further study of FurB functions. ......